Asexual Reproduction

In many simple organisms, reproduction is not a very complicated thing. It generally involves only one organism. The resulting offspring often have the exact same genetic information as the parent. This type of reproduction in which one parent is involved in the production of an identical offspring is called asexual reproduction. Asexual reproduction can take place in a variety of ways. They may include: Binary Fission: A situation in which the parent cell splits in half producing two identical cells. Fragmentation: This type of reproduction would most likely occur in molds, yeast, and mushrooms, all of which are part of the Fungi family of organisms. These organisms produce tiny filaments called Hyphae. These hyphae obtain food and nutrients from the body of other organisms in order to grow and fertilize. When a piece of hyphae breaks off and grows into a new individual, this is called fragmentation. Budding: When conditions are favorable. yeast cells can reproduce through budding. Once a copy of the genetic material is made, a bud begins to form outside the body of the yeast cell. It continues to grow larger until, eventually, it breaks away to form a new individual cell. Sporulation: Molds like to reproduce through this method. A spore is basically a reproductive cell that can grow into a new cell through mitotic cell division. Spore are stored in special spore cases until they are ready to be released. If conditions are favorable, they will grow into new individual cells. Bread mold reproduce in this manner. Regeneration: A form of asexual reproduction that take place in some invertebrates from the animal kingdom. These also produce offspring that are identical to parent. Planaria, a type of flat worm, reproduces itself by dividing in two and regenerating the missing parts. They also have the ability to regenerate injured body parts.

Spore Formation or Sporulation

In certain species of the lower bacteria, under certain circumstances, changes take place in the protoplasm which result in the formation of bodies called spores, to which the vital activities of the original bacteria are transferred. Spore formation occurs chiefly among the bacilli and in some spirilla. Its commencement in a bacterium is indicated by the appearance in the protoplasm of a minute highly refractile granule (or by a number of minute highly refractile granules scattered about throughout the protoplasm which gradually coalesce) unstained by the ordinary methods. This increases in size, and assumes a round, oval, or short rod-shaped form, always shorter but often broader than the original bacterium. In the process of spore formation the rest of

the bacterial protoplasm may remain unchanged in appearance and staining power for a considerable time (e.g. B. tetani or, on the other hand, it may soon lose its power of staining and ultimately disappear, leaving the spore in the remains of the envelope (e.g. B. anthracis). This method of spore formation is called endogenous. Bacterial spores are always non-motile. The spore may appear in the centre of the bacterium, or it may be at one extremity, or a short distance from one extremity. In structure the spore consists of a mass of protoplasm surrounded by a dense membrane. This can be demonstrated by methods which will be described, the underlying principle of which is the prolonged application of a powerful stain. The membrane is supposed to confer on the spore its characteristic feature, namely, great capacity of resistance to external influences such as heat or noxious chemicals. Koch, for instance, in one series of experiments, found that while the bacillus anthracis in the unspored form was killed by a two minutes' exposure to I per cent carbolic acid, spores of the same organism resisted an exposure of from one to fifteen days. When a spore is placed in suitable surroundings for growth it again assumes the original bacillary or spiral form. The capsule dehisces either longitudinally, or terminally, or transversely. In the last case the dehiscence may be partial, and the new individual may remain for a time attached by its ends to the hinged spore-case, or the dehiscence may be complete and the bacillus grow with a cap at each end consisting of half the spore-case. Sometimes the spore-case does not dehisce, but is simply absorbed by the developing bacterium. It is important to note that in the bacteria spore formation is rarely, if ever, to be considered as a method of multiplication. In at least the great majority of cases only one spore is formed from one bacterium, and only one bacterium in the first instance from one spore. Sporulation is to be looked upon as a resting stage of a bacterium, and is to be contrasted with the stage when active multiplication takes place. The latter is usually referred to as the vegetative stage of the bacterium. Regarding the signification of spore formation in bacteria there has been some difference of opinion. According to one view it may be regarded as representing the highest stage in the vital activity of a bacterium. There is thus an alternation between the vegetative and spore stage, the occurrence of the latter being necessary to the maintenance of the species in its greatest vitality. Such a rejuvenescence, as it were, through sporulation, is known in many algae. In support of this view there are certain facts. In many cases, for instance, spore formation only occurs at temperatures specially favourable for growth and multiplication. There is often a temperature below which, while vegetative growth still takes place, sporulation will not occur, and in the case of B. anthracis, if the organism be kept at a temperature above the limit at which it grows best, not only are no spores formed, but the species may lose the power of sporulation. Furthermore, in the case of bacteria preferring the presence of oxygen for their growth, an abundant supply of this gas may favour sporulation. Most bacteriologists are, however, of opinion that when a bacterium forms a spore, it only does so when its surroundings, especially its food supply, become unfavourable for vegetative growth ; it then remains in this condition until it is placed in more suitable surroundings. Such an occurrence would be analogous to what takes place un der similar conditions in many of the protozoa. Often sporulation can be prevented from taking place for an indefinite time if a bacterium is constantly supplied with fresh food (the other conditions of life being equal). The presence of substances excreted by the bacteria themselves plays, however/ a more important part in making the surroundings unfavourable than the mere

exhaustion of the food supply. A living spore will always develop into a vegetative form if placed in a fresh food supply. With regard to the rapid formation of spores when the conditions are favourable for vegetative growth, it must be borne in mind that in such circumstances the conditions may really very quickly become unfavourable for a continuance of growth, since not only will the food supply around the growing bacteria be rapidly exhausted, but the excretion of effete and inimical matters will be all the more rapid. We must note that the usually applied tests of a body de veloped within a bacterium being a spore are (1) its staining reaction, namely, resistance to ordinary staining fluids, but capacity of being stained by the special methods devised for the purpose ; (2) the fact that the bacterium containing the spore has higher powers of resistance against inimical conditions than a vegetative form. It is important to bear these tests in mind, as, in some of the smaller bacteria especially, it is very difficult to say whether they spore or not. There may appear in such organisms small unstained spots the significance of which it is very difficult to determine. The "missing link" is a bacterium called Acetonema longum -- a member of a little-known family of bacteria that have two membranes and respond to extreme or challenging situations by forming protective spores, a process known as sporulation. Outside of this small family, only bacteria with a single membrane have been found to sporulate. "When we started looking at Acetonema, we had no idea how unique and interesting it would turn out to be," says Grant Jensen, professor of biology at Caltech and a Howard Hughes Medical Institute investigator. "But when we imaged the sporulation process in this bacterium at high resolution, we saw that a piece of the inner membrane actually becomes the new cell's outer membrane." The finding shows that because sporulation results in a second membrane covering a cell, the common ancestor of bacteria with a single or double membrane could have been a spore-forming bacterium similar to A. longum. Jensen's group at Caltech is one of just a few in the world that uses electron cryotomography (ECT) to image biological samples. Unlike traditional electron microscopy -- for which samples must be dehydrated, embedded in plastic, sectioned, and stained -- ECT involves plunge-freezing samples so quickly that they become trapped in a near-native state within a layer of transparent, glasslike ice. A microscope can then capture high-resolution images of the sample as it is rotated, usually one degree at a time. Finally, those images can be stitched together to create threedimensional representations of a specimen such as a single bacterial cell or a virus. Postdoctoral scholar Elitza Tocheva, a member of Jensen's lab, was exploring possible projects when Jared Leadbetter, professor of environmental microbiology at Caltech, suggested the group collaborate to study a sporulating bacterium isolated from a termite gut. Leadbetter knew that the model for sporulating bacteria, Bacillus subtilis, was too thick for ECT, and this other bacterium, A. longum, happened to be relatively skinny. Tocheva quickly realized that a unique subject had fallen into her hands -- not only was A. longum thin enough to image using ECT, but it also turned out to be one of the rare spore-

forming bacteria with two membranes. The images she captured revealed the sporulation process in exquisite detail, and within the images, she noticed something interesting: in order to end up with an outer membrane after sporulation, an inner membrane had to be inverted and converted into an outer membrane. That's no small feat, given that inner and outer membranes are very different both structurally and functionally. In the case of a double-membraned bacterium, such as A. longum, the sporulation process begins with the inner membrane pinching together asymmetrically, creating a mother cell and a smaller daughter cell, all within the outer membrane. Next, the mother cell engulfs the daughter, giving the daughter an extra layer of membrane derived from the original inner membrane. The product is a spore surrounded by two membranes within the mother cell. At this point, the mother cell dies away, leaving the spore protected by those two membranes and a protein coat. When conditions improve and the spore germinates, part of its protective protein coat cracks open, allowing the new cell to outgrow, or exit, the protein coat. Unlike a single-membraned bacterium, which would at this point shed its outer membrane, the double-membraned bacterium retains it. Jensen's group found that the new outer membrane has all of the structure and function of a typical outer membrane, even though it originated as part of the mother cell's inner membrane. "When the bacterium outgrows, one of its two membranes has to be converted from an inner to an outer membrane," Tocheva says. "This is very intriguing, and we don't know how it happens." To prove that the membrane becomes converted to an outer membrane once it outgrows, the researchers had to show that Acetonema longum's outer membrane was like those seen in other double-membraned bacteria. They achieved this in a few different ways. First they showed that the outer membrane had one of the hallmarks of a true outer membrane -- the presence of an immunologically important macromolecule called lipopolysaccharide (LPS). Then they calculated the density of the membranes and found that the outer membrane was slightly denser than the inner membrane, as would be expected of a true outer membrane. Finally, they searched the genomes of about one thousand bacteria and found that A. longum had a full complement of the genes that commonly correlate with having a double membrane and having the ability to sporulate. With all of this evidence in hand, the researchers were able to say that they were indeed seeing an outer membrane that was once part of an inner membrane. "We were rewarded with this kind of rich data of a sporulating bacterium," Tocheva says. "Acetonema was just the ideal subject."

 Endospore Formation
Endospores are formed by vegetative cells in a process called sporulation. Sporulation is initiated when conditions for growth of the vegetative cells become harsh (for example, when water or an essential nutrient is limiting). Sporulation is a complex process involving as many as 200 genes. These genes are activated by an environmental trigger, causing sporulation of the cell. The steps leading to endospore formation result in the creation of a dry, metabolically inert and extremely resistant endospore from a moist, metabolically active vegetative cell. Extensive sporulation studies have been done on Bacillus subtilis, and this microbe can perform the entire sporulation process in about eight hours. Endospores can remain dormant for many years, but, when conditions allow, they can convert back to vegetative cells fairly rapidly. This process is called germination, and it involves three steps: activation, germination and outgrowth. Activation can be accomplished by heating freshly formed endospores at a high temperature. Activated spores can then be conditioned to germinate by placing them in the presence of specific nutrients. During germination, the spore becomes less resistant. This stage includes loss of calcium dipicolinate and degradation of small acid-

soluble spore proteins. The final step is outgrowth, which involves swelling due to water uptake and the synthesis of new DNA, RNA and proteins. The cell grows out of the broken spore coat and eventually resumes normal cell function. The vegetative cells continue to grow and divide until harsh environmental conditions once again trigger the sporulation process.